Methods for controlling the lysis of coagulated blood with apolipoprotein e4 phenotype

ABSTRACT

Methods for inhibiting lysis of coagulated blood and reducing risk of excessive lysis comprising administration of lysis-inhibiting amounts of apolipoprotein E4, and methods for inhibiting lysis of coagulated blood and reducing risk of excessive lysis comprising administration of a specific level of a lysis-inhibiting agent wherein the specific level is based on the apolipoprotein phenotype of an individual, are provided. Methods for enhancing lysis of coagulated blood by administration of an Apo E peptide fragment to blood containing a clot lysis agent are also provided.

RELATED APPLICATION

The present application is filed as a 371 application based onPCT/US01/31909 filed Oct. 12, 2001, which claims priority under 35U.S.C. §119 to U.S. application Ser. No. 60/240,174 filed Oct. 13, 2000.

GOVERNMENT INTERESTS

This invention was made, at least in part, with funds from the FederalGovernment, awarded through NIH grants: NO1-NS-02382, NO1-NS-02374,NO1-NS-02377, NO1-NS-02379, NO1-NS-02373, NO1-NS-02378, NO1-NS-02376,NO1-NS-02380, and NIH RO1 HL67186-01. The US government therefore hascertain acknowledged rights to the invention.

FIELD OF THE INVENTION

This invention relates to methods for inhibiting the lysis of coagulatedblood and to methods for reducing the risk of excessive lysis byadministration of a lysis-inhibiting amount of apolipoprotein E4. Theinvention further relates to methods for inhibiting the lysis ofcoagulated blood and to methods for reducing the risk of excessive lysisby the administration of a specific level of lysis-inhibiting agentwherein the specific level is based upon the apolipoprotein phenotype ofthe individual. Additionally, the invention yet further relates tomethods for enhancing the lysis of coagulated blood by theadministration of an Apo E peptide fragment to blood containing a clotlysis agent.

BACKGROUND OF THE INVENTION

Homeostasis is a term known generally to reference a procedure wherebleeding or hemorrhage is stopped by either a surgical means or throughadministration of a coagulation agent. In contrast to many thrombolytictherapies, which administer a clot lysis agent to breakdown or lysecoagulated blood associated with various ischemic diseases, the need tomaintain or enhance the coagulation of blood is also a therapeutic andbeneficial procedure.

More specifically, the administration of a clot lysis agent such astissue plasminogen activator (t-PA) within several hours following astroke is useful in the lysis of blood clots for an individual sufferingfrom an ischemic disease, such as ischemic stroke. If excessive oruncontrollable bleeding results from these therapies, however, it isbeneficial to coagulate the blood and/or stop the bleeding. As a result,the administration of a lysis-inhibiting agent may be a beneficialprocedure for treating the excessive or uncontrolled bleeding.

This beneficial process can be seen through the process of treatingindividuals suffering from hemophilia. To treat hemophilia, forinstance, many individuals are offered various blood clotting agents andfactors to cease or maintain the associated bleeding. Often theseindividuals, however, develop an insensitivity to these clottingfactors, thereby creating the need for new clotting agents to controlexcessive bleeding.

As undesirable and uncontrollable bleeding can occur in connection withmany medical disorders and ischemic diseases, for example, acute stroke,acute myocardial infarction, peripheral arterial occlusion, pulmonaryembolism, and venous thrombosis, there is a continuing need to advanceand improve current therapeutic treatments in both prophylactic andinterventional therapies.

SUMMARY OF THE INVENTION

Accordingly, it is an object of the present invention to provide novelmethods for inhibiting the lysis of coagulated blood. It is a furtherobject of the present invention to provide methods for inhibiting thelysis of coagulated blood which may be used in place of prior arttherapies.

These and additional objects are provided by the present invention. Moreparticularly, in one embodiment, the invention is directed to methodsfor inhibiting the lysis of coagulated blood, which methods compriseadministering to blood a lysis-inhibiting amount of apolipoprotein E4(Apo E4) or a therapeutic derivative thereof.

The invention is further directed to methods for reducing the risk ofexcessive blood clot lysis. These methods comprise administering toblood a lysis-inhibiting amount of Apo E4 or a therapeutic derivativethereof.

In a further embodiment, the invention is directed to methods forinhibiting the lysis of coagulated blood by administering to anindividual's blood a specific level of a lysis-inhibiting Apo E4. Thespecific level of lysis-inhibiting agent to be administered to theindividual is based upon the apolipoprotein phenotype of the individual.

In yet a further embodiment, the invention is directed to methods forreducing the risk of excessive blood clot lysis by administering to anindividual's blood a specific level of lysis-inhibiting Apo E4 whereinthe specific level is based upon the apolipoprotein phenotype of theindividual.

In another embodiment, the invention is directed to methods forenhancing the lysis of coagulated blood by administering to bloodcontaining a clot lysis agent an Apo E peptide fragment.

The methods according to the present invention are advantageous to bloodcoagulation procedures by inhibiting the lysis of coagulated bloodand/or reducing the risk of excessive blood clot lysis, particularly inindividuals at risk of exhibiting excessive or uncontrollable bleeding.Additionally, these methods are advantageous to blood coagulationprocedures by enhancing the lysis of coagulated blood in individualswith excessive blood coagulation by preventing a lysis-inhibiting effectof blood.

These and additional aspects, objects and advantages of the inventionare more fully described in the detailed description.

BRIEF DESCRIPTION OF THE DRAWINGS

The following detailed description will be more fully understood in viewof the accompanying drawings in which:

FIG. 1 depicts effects of clot lysis in vitro with respect to t-PAadministration in conjunction with increasing doses of Apo E2 or E4, asdescribed in Example 1;

FIG. 2 depicts effects of clot lysis in vitro with respect to t-PAadministration in conjunction with Apo E2 or Apo E4 as described inExample 1;

FIG. 3 a depicts a lipophilic interaction of t-PA with Apo E4 observedby Thin Layer Chromatography (TLC) as described in Example 2;

FIG. 3 b depicts a hydrophobic interaction of t-PA with Apo E4 observedby Thin Layer Chromatography (TLC) as described in Example 3; and

FIG. 4 depicts the average EC50 clot dissolution of t-PA in conjunctionwith Apo E2, Apo E3 and Apo E4 as described in Example 3.

DETAILED DESCRIPTION

The present invention is directed to methods for inhibiting the lysis ofcoagulated blood by administering to blood a lysis-inhibiting amount ofapolipoprotein E4 (Apo E4) or a therapeutic derivative thereof. Thisinvention also is directed to methods for reducing the risk ofundesirable elevated levels of blood clot lysis by administering to anindividual's blood a lysis-inhibiting amount of Apo E4 or a therapeuticderivative thereof. Also, the invention is further directed to methodsfor enhancing the lysis of coagulated blood by administering to bloodcontaining a clot lysis agent an Apo E peptide fragment.

Many current thrombolytic therapies are premised upon the administrationof a clot lysis agent to lyse coagulated blood, for example after anischemic disease is detected. In contrast to the lysis of blood,however, the frequent need to coagulate blood during such procedures orother medically related therapies is crucial when a potential forexcessive or uncontrolled bleeding exists. In addition to improving uponprevious coagulation therapy procedures, this invention is also directedto methods for inhibiting the lysis of coagulated blood and to methodsfor reducing the risk of excessive or uncontrollable bleeding byadministering a specific level of a lysis-inhibiting amount of Apo E4,wherein the specific level is based upon the apolipoprotein phenotype ofthe individual to be treated. By determining the apolipoproteinphenotype expressed by an individual, and particularly by determining ifan individual expresses Apo E4, a physician or care-giver will be ableto predict if inhibition of an administered agent will occur as a resultof the individual's inherent phenotype, and therefore more accuratelydose the lysis-inhibiting Apo E4 in an amount suitably specific for theindividual.

As used herein, “inhibiting the lysis of blood” generally refers to theinhibition, i.e., holding back, i.e., restraining of or reduction in thelysis or breakdown of coagulated blood or blood clots. As used herein,“coagulated blood” generally refers to a clot of blood such that theblood is in the form of a soft semisolid or solid mass. As used herein,“excessive lysis” generally refers to the lysis which is in excess orexceeding the normal level of lytic behavior exhibited by healthyindividuals associated with various treatments. “Clot lysis agent”generally refers to any of the known thrombolytic agents which are usedto enhance the lysis of blood clots. Known agents include, but are notlimited to, TNK-t-PA, t-PA, reteplase, streptokinase, heparin, coumadin,GIIb IIIa receptor blockers, therapeutic derivatives thereof, ormixtures thereof. Preferably, the clot lysis agent comprises tissueplasminogen activator (t-PA) or a derivative thereof.

Apolipoprotein E (Apo E) is known generally as a member of a family oflipid-associated proteins whose isoforms have been implicated as animportant modifier of several neurologic, vascular and cardiovasculardiseases. See Corder, et al. Science; 261:921–3 (1993). Apo E has threecommon isoforms—E2, E3, and E4. These monomeric isoforms combine to makesix Apo E phenotypes: E2/2, E2/3, and E2/4 (referred to as the E2category), E3/3 (E3 category), and E4/3 and E4/4 (E4 category). SeePayami, et al. JAMA; 271:1316–7 (1994); Polvikoski, et al. N Engl J Med;333:1242–1247 (1995). Apo E, and its respective isoforms, has beenlinked to outcome and survival following acute injury of the centralnervous system, as well as peripheral cardiovascular system. Thepresence of an E4 allele has been associated with a poor outcomefollowing severe head trauma and with poorer survival in patients withan intracerebral hemorrhage (ICH). See Alberts et al, Stoke 27: 183.,1986; Freidman et al, Neurology, 52: 244–248., 1999. The E4 isoforms,E4/3 and E4/4, referred to herein as Apo E4, are employed in the methodsof the invention. Therapeutic derivatives can also be administered.

As used herein, “therapeutic derivative” generally refers to an Apo E4fragment, for example fragments of Apo E4 having the lysis-inhibitingactivity, or chemical or structural analogue exhibiting thelysis-inhibiting activity. “Lysis inhibiting amount” generally refers toa quantitative amount of Apo E4 which inhibits lysis activity of blood,with or without a conventional clot lysis agent. Suitable doses of ApoE4, or its therapeutic derivative, necessary to inhibit the lyticactivity of a naturally occurring or an administered clot lysis agentsuch as t-PA will vary depending, inter alia, on physiologicalcharacteristics of the individual to be treated, including, but notlimited to the Apo E phenotype of the individual, and on the givenmedical condition of the individual. However, typically, the Apo E4 orderivative is administered in an amount of about equal molarstoicheometry with the clot lysis agent to provide the lytic inhibitingactivity. The equimolar amount is based on the amount of clot lysisagent administered, if any, and the amount of clot lysis agent inherentin the blood. For example, human blood typically contains about 0.0009mg/l t-Pa.

The dosages of conventional clot lysis agents are generally based uponthe individual's physiological characteristics and given medicalcondition. Typically a patient can be administered, for example, a rangeof t-PA of from about 0.1–10.0 mg/kg of blood. A standard therapeuticdose of t-PA physiologically appropriate for thrombolytic therapy to beadministered intravenously to blood over a one hour period is 0.9 mg/kgof blood.

“Ischemic disease” generally refers to a medical condition causing orresulting in a decrease in blood supply to a bodily organ, tissue, orlocation due to constriction or obstruction of blood vessels. Examplesof ischemic diseases comprise myocardial infarction, unstable angina,coronary artery thrombus, and peripheral vascular disease. “Peripheralvascular diseases” typically refers to, inter alia, occlusions,retinopathy, and organ embolisms. “Organ embolism” typically refers tothe obstruction or occlusion of a blood vessel by an embolus within anorgan system, for example, such as a pulmonary embolism. In treatingthese and similar ischemic diseases, thrombolytic therapy in the form ofa clot lysis agent is often administered to enhance the lysis ofcoagulated blood. In addition to the need to enhance the lysis of blood,often many medical procedures and thrombolytic therapies may result inexcessive bleeding, thereby requiring the administration of acoagulating agent or factor. For instance, some diseases and proceduresthat may be effected by such thrombolytic therapy and thereby requirethe administration of a procoagulant include, surgery, vascular surgery,grafts, organ transplants, limb reattachment, trauma pregnancy, childbirth, post partum hemophilia, and normal vaginal delivery withexcessive post partum bleeding. The methods for inhibiting the lysis ofcoagulated blood according to the present invention may be employed intreatment of any such diseases or conditions. Additionally, individualssuffering from hemophilia typically require the administration ofclotting agents such as Factor VIII to coagulate the blood. Suchprocedures can also benefit by administering Apo E4 as a clotlysis-inhibiting agent in conjunction with the conventional Factor VIIItherapeutic procedures.

In one embodiment, a lysis-inhibiting amount of Apo E4 is administeredto an individual with an ischemic disease to inhibit the lysis ofcoagulated blood associated with the disease.

In a specific embodiment, methods for reducing the risk of excessiveblood clot lysis can be administered, particularly in individuals atrisk of undesirable or excessive bleeding. These methods compriseadministering to blood a lysis-inhibiting amount of Apo E4 or atherapeutic derivative thereof.

In a further embodiment, a specific level of a lysis-inhibiting amountof Apo E4 is administered wherein the specific level is based upon theapolipoprotein phenotype of the individual. In determining the specificlevel of agent to be administered to the individual, the apolipoproteinphenotype of the individual will help the physician or care-giverconclude how much lysis-inhibiting Apo E4 will be required to obtaindesired results. For example, a lesser amount of lysis-inhibiting Apo E4will typically be required if the individual has an Apo E4 phenotype,due to the clot lysis inhibiting properties exhibited by Apo E4.Preferably, the individual's apolipoprotein phenotype is determinedprior to administering the specific level of lysis-inhibiting agent.Methods for determining apolipoprotein phenotype from blood or serumsamples are known in the art and may be employed.

In another specific embodiment, the lysis-inhibiting amount of Apo E4 isadministered to an individual with post surgical complications ofocclusion or clotting to inhibit the lysis of blood clots associatedwith the ischemic disease. Furthermore, a specific level of alysis-inhibiting amount of Apo E4 can be administered to an individualbased upon the apolipoprotein phenotype of the individual to inhibit thelysis of coagulated blood associated with post surgical complications ofocclusion or clotting.

In yet another embodiment, an Apo E peptide fragment is administered toblood containing a clot lysis agent to enhance the lysis of coagulatedblood via a competitive antagonist procedure. In administering the Apo Epeptide fragment, it is possible to prevent the natural lysis-inhibitingeffect of Apo E4 within blood. For example, an Apo E fragment which hasfunctionality for binding with a clot lysis agent but does not exhibitthe lysis inhibiting effect of Apo E4 can be administered. As thefragment binds to the clot lysis agent, for example t-PA, it preventsbinding of lysis inhibiting Apo E4. The process of competitiveinhibition of the inhibition can be used to prevent the naturalinhibiting effect of Apo E4 in blood by administering a peptide fragmentof Apo E with an associated binding site but without thelysis-inhibiting characteristics. Upon administering this peptidefragment of Apo E into the blood containing Apo E4 and a lysis agent,the structures compete for the same binding site location, therebyallowing some fragments of Apo E to bind to the clot lysis agent priorto Apo E4 binding. As a result, the Apo E peptide fragments protect theblood from receiving the lysis-inhibiting effects associated with theApo E4 peptides by connecting to the binding site and thereby blockingthe Apo E4 interaction. Derivatives through combinational chemistry ofthe structure, ionic, or electron density map of the Apo E peptide canalso be employed as the Apo E fragment to bind to a lysis agent such ast-PA and prevent the lysis-inhibiting effect of the larger molecules ofApo E.

The following example is provided to illustrate the methods and variousembodiments of the present invention. While the example below utilizest-PA as the clot lysis agent for demonstrating the lysis-inhibitingeffects of Apo E4, similar results can be obtained with other clot lysisagents as mentioned above. The use of similar clot lysis agents will beapparent to one of ordinary skill in the art and are within the scope ofthe claims.

EXAMPLE 1

In this example, the lysis-inhibiting effect of Apo E4 is demonstratedin vitro. There are various methods for determining the function, actionand or kinetics of t-PA. These include but are not limited to: measuringfibrin degradation fragments, clot lysis, clot weight, fluid evolutionweight, clot times and others. Those experienced in the art will befamiliar with these and other methods for assaying t-PA activity or clotdegradation/formation etc. The data from this study consists of ameasurement of clot dissolution or lysis by measuring the amount ofliquid liberated from the blood sample. This is an index of clot lysisand the clot lysis is stimulated by t-PA.

Typically, two mls of fresh human blood are drawn and added topre-weighed vials containing known amounts of t-PA with or without ApoE4. The blood is mixed and allowed to equilibrate for 40 minutes. Thevials are centrifuged at 1 g for 15 minutes and the supernatantdecanted. Clot weight and solution weight is determined and used tomeasure clot degradation or dissolution. Increasing supernatant volumesindicate increased clot degradation or increasing clot dissolution.

FIG. 1 depicts the effect of clot lysis on the blood from twoindividuals with respect to t-PA administration in conjunction withincreased dosages of Apo E2 or Apo E4. The y-axis demonstrates in grams(g) the amount of clot lysis or dissolution measured. The administrationof t-PA to individual A's blood at a level of 1.0 mg/l blood inconjunction with increasing amounts of Apo E4 shows a dose dependentdecrease in clot dissolution. In the top two lines representingindividual B's blood, there was no significant change in clotdissolution caused by E4 after administrating t-PA at a level of 0.5mg/l blood. This is believed to be due to administration of aspirin tothe individual prior to sampling. N=3 for each.

FIG. 2 depicts the effect of clot lysis on additional blood fromindividual B with respect to t-PA administration in combination with ApoE2 or Apo E4. The y-axis demonstrates in grams (g) the amount of clotlysis or dissolution measured. The administration of t-PA alone at alevel of 0.5 mg/l blood shows that the level of clot lysis wassubstantially complete at 2.5 g. Upon administration of t-PA inconjunction with Apo E2, no significant change in lytic activity wasrecognized. The amount of clot lysis was significantly inhibited,however, upon the administration of t-PA in conjunction with Apo E4. N=3for each.

EXAMPLE 2

In this example, the lipophilic interaction of t-PA with Apo E4 isdemonstrated through Thin Layer Chromatography (TLC). There are numerousanalytical methods for measuring molecular interactions. These are knownto those experienced in the art and include, but are not limited to;Thin Layer Chromatography (TLC), Paper Chromatography, electrophoresis,diffraction methods, Nuclear Magnetic Resonance, and other methods.

A known concentration of t-PA in a first prepared solution is applied toa TLC plate and allowed to dry to measure molecular interaction. In asecond solution equal molar amounts of t-PA and Apo E4 are employed andthe solution is similarly applied to a second TLC plate and allowed todry.

The TLC plates are put into migration chambers containingchloroform:ethanol in a 3:1 or 1:1 ratio and allowed to migrate untilthe solvent front approaches 1 cm from the top of the plate. The platesare removed from the chambers and allowed to dry in the fume cupboards.The plates can be read with ninhydrin or uv light quenching. Themigration distances are marked and rf values reported. The plate towhich the first solution containing only t-PA is applied is used as areference and the percent increase in migration exhibited by the secondsolution is set forth in FIG. 3 a, N=3 for each.

The results demonstrate that Apo E4 reacts and uniquely binds with t-PAunder the given conditions, presumably through the lipophilic proteinproperties although the inventors do not intend to be bound by thistheory.

EXAMPLE 3

In this example, using an in vitro clot degradation method, the lysisinhibiting effect of Apo E4 on t-PA is further demonstrated. Theinhibition of clot dissolution by Apo E4 is statistically significant.

The ability of Apo E isoproteins to modulate t-PA induced clot lysis isassessed using an in vitro clot assay system. This system uses thedecrease in clot formation in the presence of t-PA to approximate theamount of clot lysis. Blood samples are obtained from 18 volunteers anddivided into three Apo E genotypes (6 patients in each group): E2(E2/E2, E2/E3, E2/E4), E3 (E3/E3) and E4 (E3/E4, E4/E4). Clot lysis inthe presence of varying concentrations of t-PA (0 to 4 μg/ml), isassessed in the presence or absence of supplemental Apo E2, E3 or E4(9.8 μg/ml) for each patient genotype. The results are expressed asEC50s, which are the effective concentrations of t-PA required toachieve 50% lysis of the clot.

As shown in FIG. 4, t-PA induced clot lysis is significantly (P±0.0001)enhanced by supplementation with Apo E2 (EC50 of 0.20±0.06 μg/ml) ascompared to t-PA alone (0.72±0.19). When Apo E4 is supplemented to theclot lysis assay, there is a significant (P<0.05) inhibition of clotlysis (EC50 of 0.98±0.23 μg/ml) as compared to t-PA alone (0.72±0.19),but there is no significant change in t-PA induced clot lysis caused byApo. E3. Examining the relationship between patient genotypes and clotlysis, there is a significant increase in clot lysis for all groups withApo E2 supplementation, and a non-significant trend for the Apo E4patient group to have decreased clot lysis with Apo E3 and E4supplementation. The results demonstrate that the t-PA is workingthrough a clot degradation mechanism.

Using thin layer chromatography (TLC), as defined in Example 2, Apo E2and E4 interacted chemically with t-PA, and Apo E3 either did notinteract with t-PA or that said interaction is distinctly different thanthe interaction seen with E2 and E4 (as seen if FIG. 3 b). The resultsdemonstrate that Apo E4 reacts and uniquely binds with t-PA under thegiven conditions, through hydrophobic domains although the inventors donot intend to be bound by this theory.

The specific embodiments and examples set forth above are provided forillustrative purposes only and are not intended to limit the scope ofthe following claims. Additional embodiments of the invention andadvantages provided thereby will be apparent to one of ordinary skill inthe art and are within the scope of the claims.

1. A method for inhibiting lysis of coagulated blood, comprisingadministering to blood a lysis-inhibiting amount of apolipoprotein E4(Apo E4) or a therapeutic derivative thereof.
 2. The method as definedby claim 1, wherein the blood comprises a clot lysis agent.
 3. Themethod as defined by claim 2, wherein the clot lysis agent comprisestissue plasminogen activator (t-PA).
 4. The method as defined by claim2, wherein the clot lysis agent comprises a t-PA derivative.
 5. Themethod as defined by claim 2, wherein the clot lysis agent comprisesTNK-t-PA, t-PA, reteplase, streptokinase, heparin, coumadin, GIIb IIIareceptor blockers, therapeutic derivatives thereof, or mixtures thereof.6. The method as defined by claim 1, wherein a therapeutic derivativecomprising fragments of Apo E4 having lysis-inhibiting activity isadministered.
 7. The method as defined by claim 1, wherein thelysis-inhibiting amount of Apo E4 is administered to an individual withan ischemic disease.
 8. The method as defined by claim 7, wherein theischemic disease comprises myocardial infarction, unstable angina,coronary artery thrombus, or peripheral vascular disease.
 9. The methodas defined by claim 8, wherein the peripheral vascular disease comprisesocclusion, retinopathy, or organ embolism.
 10. The method as defined byclaim 9, wherein the organ embolism comprises pulmonary embolism. 11.The method as defined by claim 1, wherein the lysis-inhibiting amount ofApo E4 is administered to an individual with post surgical complicationsof occlusion or clotting.
 12. A method for reducing risk of excessiveblood clot lysis, comprising administering to blood a lysis-inhibitingamount of Apo E4 or a therapeutic derivative thereof.
 13. A method forinhibiting lysis of coagulated blood, comprising administering to anindividual's blood a specific level of a lysis-inhibiting amount of ApoE4, wherein the specific level is based upon an apolipoprotein phenotypeof the individual.
 14. The method as defined by claim 13, wherein theapolipoprotein phenotype is Apo E4.
 15. The method as defined by claim13, wherein the blood comprises a clot lysis agent.
 16. The method asdefined by claim 15, wherein the clot lysis agent comprises tissueplasminogen activator (t-PA).
 17. The method as defined by claim 15,wherein the clot lysis agent comprises a t-PA derivative.
 18. The methodas defined by claim 15, wherein the clot lysis agent comprises TNK-t-PA,t-PA, reteplase, streptokinase, heparin, coumadin, GIIb IIIa receptorblockers, therapeutic derivatives thereof, or mixtures thereof.
 19. Themethod as defined by claim 13, wherein a therapeutic derivativecomprising fragments of Apo E4 having lysis-inhibiting activity isadministered.
 20. The method as defined by claim 13, wherein theindividual has an ischemic disease.
 21. The method as defined by claim20, wherein the ischemic disease comprises myocardial infarction,unstable angina, coronary artery thrombus, or peripheral vasculardisease.
 22. The method as defined by claim 21, wherein the peripheralvascular disease comprises occlusion, retinopathy, or organ embolism.23. The method as defined by claim 22, wherein the organ embolismcomprises pulmonary embolism.
 24. The method as defined by claim 13,wherein the lysis-inhibiting amount is administered to an individualwith post surgical complications of occlusion or clotting.
 25. Themethod as defined by claim 13, comprising the additional step ofdetermining the individual's apolipoprotein phenotype prior toadministering the specific level of lysis-inhibiting Apo E4.
 26. Amethod for reducing risk of excessive blood clot lysis, comprisingadministering to an individual's blood a specific level of alysis-inhibiting amount of Apo E4, wherein the specific level is basedupon an apolipoprotein phenotype of the individual.
 27. The method asdefined by claim 26, comprising the additional step of determining theindividual's apolipoprotein phenotype prior to administering thespecific level of lysis-inhibiting Apo E4.